The symptoms of obstructive lung diseases, which include airways hyperresponsiveness, reversible airways obstruction, chest tightness, dyspnea, mucus hypersecretion and cough, are in large part the result of an excessive and/ or inappropriate activation of the vagal afferent nerves innervating the airways and lungs. Understanding the mechanisms by which these sensory nerves are activated in health and disease and the reflexes and sensations evoked as a consequence of their activation have been longstanding goals of the research carried out in our laboratories. Visceral afferent nerves often rely on specialized chemosensory signaling mechanisms at their nerve terminals to transduce mucosal irritation. We recently described a chemosensory transduction pathway in the airways mucosa that relies on the transmitter acetylcholine and the nicotinic subclass of acetylcholine receptors (nAChRs) to initiate changes in breathing pattern in response to mucosal irritation. The involvement of nAChRs is noteworthy, as their activity is inappropriately and excessively recruited in the airways of patients exposed to cigarette smoke or the nicotine-containing vapors of eCigarettes. The central hypothesis of this research proposal is that nAChRs play essential roles in transducing reflexes initiated endogenous irritants acting on subsets of vagal sensory nerves, and modulate airway defensive reflexes both peripherally and centrally. We also hypothesize that the regulatory functions of nAChRs are corrupted by chronic mucosal irritation associated with asthma and COPD, and by smoking. Studies proposed herein aim to: 1) characterize the vagal afferent nerve subtypes responsible for nAChR- dependent exacerbations of obstructive lung diseases and the nAChR subtypes involved; 2a) determine the transduction mechanisms for nAChR-dependent coughing and other airway defensive reflexes both at the peripheral nerve terminals and centrally at the termination sites of the neural crest C-fibers that are selectively stimulated through nAChR activation; 2b) once the mechanisms for nAChR-dependent reflexes are established in healthy animals, we will evaluate changes in transduction mechanisms following chronic nicotine administration and during allergen-induced cough hypersensitivity; and 3) building upon our intriguing recent discovery of ?7 nAChR subtype dependent inhibition of evoked coughing through central sites of action, we will determine the CNS effects of nAChR activation on airway defensive reflexes, and how these central regulatory pathways are altered by chronic nicotine administration and during cough hypersensitivity. For all of these studies, we will utilize techniques that are unique to our laboratories, including single cell recording and molecular approaches, reflex physiological recordings and in vivo imaging of nAChR occupancy. We anticipate that the results of these proposed studies will reveal the novel and essential roles of nAChRs in transducing mucosal irritation in the airways of patients with chronic diseases of the airways and lungs.